Cell type and dynamic state govern genetic regulation of gene expression in heterogeneous differentiating cultures.

Identifying the molecular effects of human genetic variation across cellular contexts is crucial for understanding the mechanisms underlying disease-associated loci, yet many cell types and developmental stages remain underexplored. Here, we harnessed the potential of heterogeneous differentiating cultures (HDCs), an in vitro system in which pluripotent cells asynchronously differentiate into a broad spectrum of cell types. We generated HDCs for 53 human donors and collected single-cell RNA sequencing data from over 900,000 cells.

Oxygen-induced stress reveals context-specific gene regulatory effects in human brain organoids.

The interaction between genetic variants and environmental stressors is key to understanding the mechanisms underlying neurological diseases. In this study, we used human brain organoids to explore how varying oxygen levels expose context-dependent gene regulatory effects. By subjecting a genetically diverse panel of 21 brain organoids to hypoxic and hyperoxic conditions, we identified thousands of gene regulatory changes that are undetectable under baseline conditions, with 1,745 trait-associated genes showing regulatory effects only in response to oxygen stress.

WIPI2b recruitment to phagophores and ATG16L1 binding are regulated by ULK1 phosphorylation.

One of the key events in autophagy is the formation of a double-membrane phagophore, and many regulatory mechanisms underpinning this remain under investigation. WIPI2b is among the first proteins to be recruited to the phagophore and is essential for stimulating autophagy flux by recruiting the ATG12-ATG5-ATG16L1 complex, driving LC3 and GABARAP lipidation. Here, we set out to investigate how WIPI2b function is regulated by phosphorylation.

Cell-type and dynamic state govern genetic regulation of gene expression in heterogeneous differentiating cultures.

Identifying the molecular effects of human genetic variation across cellular contexts is crucial for understanding the mechanisms underlying disease-associated loci, yet many cell-types and developmental stages remain underexplored. Here we harnessed the potential of heterogeneous differentiating cultures ( ), an system in which pluripotent cells asynchronously differentiate into a broad spectrum of cell-types. We generated HDCs for 53 human donors and collected single-cell RNA-sequencing data from over 900,000 cells.

Steroid receptor coactivators in Treg and Th17 cell biology and function.

Steroid receptor coactivators (SRCs) are master regulators of transcription that play key roles in human physiology and pathology. SRCs are particularly important for the regulation of the immune system with major roles in lymphocyte fate determination and function, macrophage activity, regulation of nuclear factor κB (NF-κB) transcriptional activity and other immune system biology. The three members of the p160 SRC family comprise a network of immune-regulatory proteins that can function independently or act in synergy with each other, and compensate for - or moderate - the activity of other SRCs.

Genetic regulatory effects in response to a high-cholesterol, high-fat diet in baboons.

Steady-state expression quantitative trait loci (eQTLs) explain only a fraction of disease-associated loci identified through genome-wide association studies (GWASs), while eQTLs involved in gene-by-environment (GxE) interactions have rarely been characterized in humans due to experimental challenges. Using a baboon model, we found hundreds of eQTLs that emerge in adipose, liver, and muscle after prolonged exposure to high dietary fat and cholesterol. Diet-responsive eQTLs exhibit genomic localization and genic features that are distinct from steady-state eQTLs.

Effects of Aircraft Noise on Sleep: Federal Aviation Administration National Sleep Study Protocol.

Aircraft noise can disrupt sleep and impair recuperation. The last U.S.

Induced Pluripotent Stem Cells in Disease Biology and the Evidence for Their In Vitro Utility.

Many human phenotypes are impossible to recapitulate in model organisms or immortalized human cell lines. Induced pluripotent stem cells (iPSCs) offer a way to study disease mechanisms in a variety of differentiated cell types while circumventing ethical and practical issues associated with finite tissue sources and postmortem states. Here, we discuss the broad utility of iPSCs in genetic medicine and describe how they are being used to study musculoskeletal, pulmonary, neurologic, and cardiac phenotypes.

The relationship between regulatory changes in cis and trans and the evolution of gene expression in humans and chimpanzees.

Background: Comparative gene expression studies in apes are fundamentally limited by the challenges associated with sampling across different tissues. Here, we used single-cell RNA sequencing of embryoid bodies to collect transcriptomic data from over 70 cell types in three humans and three chimpanzees.

Results: We find hundreds of genes whose regulation is conserved across cell types, as well as genes whose regulation likely evolves under directional selection in one or a handful of cell types.

Genetic regulatory effects in response to a high cholesterol, high fat diet in baboons.

Steady-state expression quantitative trait loci (eQTLs) explain only a fraction of disease-associated loci identified through genome-wide association studies (GWAS), while eQTLs involved in gene-by-environment (GxE) interactions have rarely been characterized in humans due to experimental challenges. Using a baboon model, we found hundreds of eQTLs that emerge in adipose, liver, and muscle after prolonged exposure to high dietary fat and cholesterol. Diet-responsive eQTLs exhibit genomic localization and genic features that are distinct from steady-state eQTLs.

Guided Differentiation of Pluripotent Stem Cells for Cardiac Cell Diversity.

We have developed a guided differentiation protocol for induced pluripotent stem cells (iPSCs) that rapidly generates a temporally and functionally diverse set of cardiac-relevant cell types. By leveraging techniques used in embryoid body and cardiac organoid generation, we produce both progenitor and terminal cardiac cell types concomitantly in just 10 days. Our results show that guided differentiation generates functionally relevant cardiac cell types that closely align with the transcriptional profiles of cells from differentiation time-course collections, mature cardiac organoids, and heart tissue.

Steroid receptor coactivator 3 is a key modulator of regulatory T cell-mediated tumor evasion.

Steroid receptor coactivator 3 (SRC-3) is most strongly expressed in regulatory T cells (Tregs) and B cells, suggesting that it plays an important role in the regulation of Treg function. Using an aggressive E0771 mouse breast cell line syngeneic immune-intact murine model, we observed that breast tumors were "permanently eradicated" in a genetically engineered tamoxifen-inducible Treg-cell-specific SRC-3 knockout (KO) female mouse that does not possess a systemic autoimmune pathological phenotype. A similar eradication of tumor was noted in a syngeneic model of prostate cancer.

Identifying a selective inhibitor of autophagy that targets ATG12-ATG3 protein-protein interaction.

Macroautophagy/autophagy is a catabolic process by which cytosolic content is engulfed, degraded and recycled. It has been implicated as a critical pathway in advanced stages of cancer, as it maintains tumor cell homeostasis and continuous growth by nourishing hypoxic or nutrient-starved tumors. Autophagy also supports alternative cellular trafficking pathways, providing a mechanism of non-canonical secretion of inflammatory cytokines.

Steroid Receptor Coactivator-3 is a Key Modulator of Regulatory T Cell-Mediated Tumor Evasion.

Unlabelled: Steroid receptor coactivator 3 (SRC-3) is most strongly expressed in regulatory T cells (Tregs) and B cells, suggesting that it plays an important role in the regulation of Treg function. Using an aggressive E0771 mouse breast cell line syngeneic immune-intact murine model, we observed that breast tumors were 'permanently eradicated' in a genetically engineered tamoxifen-inducible Treg-cell specific SRC-3 knockout (KO) female mouse that does not possess a systemic autoimmune pathological phenotype. A similar eradication of tumor was noted in a syngeneic model of prostate cancer.

Steroid receptor coactivators - their role in immunity.

Steroid Receptor Coactivators (SRCs) are essential regulators of transcription with a wide range of impact on human physiology and pathology. In immunology, SRCs play multiple roles; they are involved in the regulation of nuclear factor-κB (NF-κB), macrophage (MΦ) activity, lymphoid cells proliferation, development and function, to name just a few. The three SRC family members, SRC-1, SRC-2 and SRC-3, can exert their immunological function either in an independent manner or act in synergy with each other.

Evolutionary insights into primate skeletal gene regulation using a comparative cell culture model.

The evolution of complex skeletal traits in primates was likely influenced by both genetic and environmental factors. Because skeletal tissues are notoriously challenging to study using functional genomic approaches, they remain poorly characterized even in humans, let alone across multiple species. The challenges involved in obtaining functional genomic data from the skeleton, combined with the difficulty of obtaining such tissues from nonhuman apes, motivated us to consider an alternative in vitro system with which to comparatively study gene regulation in skeletal cell types.

Human embryoid bodies as a novel system for genomic studies of functionally diverse cell types.

Practically all studies of gene expression in humans to date have been performed in a relatively small number of adult tissues. Gene regulation is highly dynamic and context-dependent. In order to better understand the connection between gene regulation and complex phenotypes, including disease, we need to be able to study gene expression in more cell types, tissues, and states that are relevant to human phenotypes.

Single-cell sequencing reveals lineage-specific dynamic genetic regulation of gene expression during human cardiomyocyte differentiation.

Dynamic and temporally specific gene regulatory changes may underlie unexplained genetic associations with complex disease. During a dynamic process such as cellular differentiation, the overall cell type composition of a tissue (or an in vitro culture) and the gene regulatory profile of each cell can both experience significant changes over time. To identify these dynamic effects in high resolution, we collected single-cell RNA-sequencing data over a differentiation time course from induced pluripotent stem cells to cardiomyocytes, sampled at 7 unique time points in 19 human cell lines.

Benchmarking sequencing methods and tools that facilitate the study of alternative polyadenylation.

Background: Alternative cleavage and polyadenylation (APA), an RNA processing event, occurs in over 70% of human protein-coding genes. APA results in mRNA transcripts with distinct 3' ends. Most APA occurs within 3' UTRs, which harbor regulatory elements that can impact mRNA stability, translation, and localization.

Functional dynamic genetic effects on gene regulation are specific to particular cell types and environmental conditions.

Genetic effects on gene expression and splicing can be modulated by cellular and environmental factors; yet interactions between genotypes, cell type, and treatment have not been comprehensively studied together. We used an induced pluripotent stem cell system to study multiple cell types derived from the same individuals and exposed them to a large panel of treatments. Cellular responses involved different genes and pathways for gene expression and splicing and were highly variable across contexts.

A genome-scale CRISPR Cas9 dropout screen identifies synthetically lethal targets in SRC-3 inhibited cancer cells.

Steroid receptor coactivator 3 (SRC-3/NCoA3/AIB1), is a key regulator of gene transcription and it plays a central role in breast cancer (BC) tumorigenesis, making it a potential therapeutic target. Beyond its function as an important regulator of estrogen receptor transcriptional activity, SRC-3 also functions as a coactivator for a wide range of other transcription factors, suggesting SRC-3 inhibition can be beneficial in hormone-independent cancers as well. The recent discovery of a potent SRC-3 small molecule inhibitor, SI-2, enabled the further development of additional related compounds.

Divergence in alternative polyadenylation contributes to gene regulatory differences between humans and chimpanzees.

While comparative functional genomic studies have shown that inter-species differences in gene expression can be explained by corresponding inter-species differences in genetic and epigenetic regulatory mechanisms, co-transcriptional mechanisms, such as alternative polyadenylation (APA), have received little attention. We characterized APA in lymphoblastoid cell lines from six humans and six chimpanzees by identifying and estimating the usage for 44,432 polyadenylation sites (PAS) in 9518 genes. Although APA is largely conserved, 1705 genes showed significantly different PAS usage (FDR 0.

Drug Combination in Cancer Treatment-From Cocktails to Conjugated Combinations.

It is well recognized today that anticancer drugs often are most effective when used in combination. However, the establishment of chemotherapy as key modality in clinical oncology began with sporadic discoveries of chemicals that showed antiproliferative properties and which as a first attempt were used as single agents. In this review we describe the development of chemotherapy from its origins as a single drug treatment with cytotoxic agents to polydrug therapy that includes targeted drugs.